1. Field of the Invention
This invention relates to the separation of enantiomers. This invention more particularly relates to a chiral selector for separation of enantiomers. This invention especially relates to a chiral selector useful, for example, as a chiral stationary phase (CSP) in the liquid chromatographic (HPLC) separation of enantiomers of underivatized beta-amino alcohol compounds.
2. Description of the Prior Art
Stereoisomers are those molecules which differ from each other only in the way their atoms are oriented in space. Stereoisomers are generally classified as diastereomers or enantiomers; the latter embracing those which are mirror-images of each other, the former being those which are not. The particular arrangement of atoms that characterize a particular stereoisomer is known as its optical configuration, specified by known sequencing rules as, for example, either + or - (also D or L) and/or R or S.
Though differing only in orientation, the practical effects of stereoisomerism are important. For example, the biological and pharmaceutical activities of many compounds are strongly influenced by the particular configuration involved. Indeed, many compounds are only of widespread utility when provided in a given stereoisomeric form. Living organisms usually produce only one enantiomer of a pair. Only (-)-2-methyl-1-butanol is formed in yeast fermentation of starches; only (+)-lactic acid is formed in the contraction of muscle; fruit juices contain only (-)-malic acid and only (-)-quinine is obtained from a cinchona tree. In biological systems, stereochemical specificity is the rule rather than the exception, since the catalytic enzymes which are so important in such systems are optically active. The sugar (+)-glucose plays an important rule in animal metabolism and is the basic raw material in the fermentation industry; yet (-)-glucose is not metabolized by animals or fermented by yeasts. The mold Penicillium glaucum will only consume the (+)-enantiomer of an enantiomeric mixture of tartaric acid leaving (-)-tartaric acid intact. Only one stereoisomer of chloromycetin is an antibiotic. Not only does (+)-ephedrine not have any drug activity, but it interferes with the drug activity of its enantiomer. (-)-Carvone provides oil of spearmint with its distinctive odor while its enantiomer, (+)-carvone has the essence of caraway. Hence, it is desirable and often essential to separate stereoisomers to provide the useful version of an optically active chemical compound.
When diastereomers are involved, separation is generally not a significant problem because diastereomers have different physical properties, such as melting points, boiling points, solubilities in a given solvent, densities, refractive indices, etc. Hence, diastereomers may normally be separated from each other by conventional methods, such as fractional distillation, fractional crystallization, or chromatography.
Enantiomers, on the other hand, present a special problem because their physical properties are identical. They cannot as a rule--especially when in the form of a racemic mixture--be separated by ordinary methods: not by fractional distillation, because their boiling points are identical; not by conventional fractional crystallization, because (unless the solvent is optically active) their solubilities are identical; not by conventional chromatography because (unless the adsorbent is optically active) they are held equally onto the adsorbent. The problem of separating enantiomers is further exacerbated by the fact that conventional synthetic techniques almost always produce a mixture of enantiomers. When the mixture comprises equal amounts of enantiomers having different optical configurations, it is called a racemate or a racemic modification; and separation of the racemate into its respective enantiomers--this separation being generally known as a resolution--is, therefore, a process of considerable importance.
Various techniques for separating enantiomers are known. Most, however, are directed to small analytical quantities, meaning that, other drawbacks aside, when applied to preparative scale amounts (the milligram to kilogram range) a loss of resolution occurs. Hand separation--the oldest method of resolution--is not only impractical but can almost never be used since racemates seldom form mixtures of crystals recognizable as mirror-images.
Another method, known as an indirect separation, involves the conversion of a mixture of enantiomers--the racemate--into a mixture of diastereomers. The conversion is accomplished by reacting the enantiomers with an optically pure chiral derivatizing agent. The resultant diastereomers are separated from each other by taking advantage of their different physical properties. Once separated, by, for example, fractional crystallization, or more commonly, chromatography, the diastereomers are reconverted back into the corresponding enantiomers, which are now optically pure. Though achieving the requisite separation, the indirect method suffers in that it is time-consuming and can require large quantities of optically pure derivatizing agent which can be expensive and is oftentimes not recoverable. Moreover, the dederivatization step may itself result in racemization thus defeating the purpose of the separation earlier achieved.
A more current method which avoids some of the drawbacks attendant the indirect method is known as the direct method of separation. The direct method, much like the indirect method, involves the formation of a diastereomeric species. However, unlike the indirect method, this species is transient, with the stability of one species differing from the other.
In one application of the direct method, the mixture of enantiomers is allowed to interact with a chiral stationary phase (CSP), which, for example, could reside in a chromatographic column. The enantiomer that interacts more strongly with the chiral stationary phase than the other will have a longer residence time on the chiral stationary phase and hence a separation will occur. When the mode of interaction with the chiral stationary phase can be characterized, the elution order may be predicted. Examples of chiral stationary phases include those based on (L)-N-(3,5-dinitrobenzoyl)leucine, which is useful in separating enantiomers of N-aryl derivatized amino acids and esters and those based on (L)-N-(1-naphthyl)leucine which has been used to effectively separate N-(3,5-dinitrobenzoyl) derivatized amino compounds. HPLC columns packed with silica-bonded CSPs of a variety of pi-electron acceptors and pi-electron donors, including derivatives of phenylglycine, leucine, naphthyl-alanine and naphthylleucine are commercially available from Regis Chemical Company, Morton Grove, Ill.
In another application of the direct method, disclosed in U.S. Pat. No. 5,080,795 to Pirkle et al., enantiomers of such compounds as amino acids, amino esters, sulfonides, alcohols, amines, sulfonic acids or derivatives thereof are separated by means of a liquid membrane containing a chiral carrier, such as derivatized amino acid, (S)-N-(1-naphthyl)leucine octadecyl ester. The chiral carrier is capable of forming a stable complex with one of the enantiomers. The liquid membrane is located on one side of a semi-permeable barrier, and the mixture of enantiomers is located on the other side of the liquid membrane. The liquid membrane containing the chiral carrier impregnates the semi-permeable barrier under conditions effective to permit or cause a stable complex between the chiral carrier and one of the enantiomers to form in the liquid membrane. The liquid membrane containing the stable complex is passed to a second location where the conditions are effective to dissociate the stable complex and the recovery of the complex-forming enantiomer. In one embodiment of this application, a hollow-fiber membrane is employed as the semi-permeable barrier.
It is widely recognized that stereoisomers of pharmaceutical agents may have drastically different pharmacological potencies or actions. For example, the so-called .beta.-blockers, widely used in the treatment of angina pectoris and hypertension, differ considerably in the physiological responses they elicit. Typically, the (S) enantiomers are 50-500 fold more active than their antipodes and may differ also in the nature of the elicited responses. .beta.-blockers are adrenergic blocking agents capable of blocking nerve impulses to special sites (beta acceptors) in the cerebellum in order to reduce the heartbeat rate and the force of heart contractions. Owing to their importance, many potential .beta.-blockers have been developed and tested, and a number are now marketed. Known .beta.-blockers include compounds identified as metoprolol, oxprenolol, propanolol, pindolol, pronethalol and bufuralol. The common aspect of the compounds is that they all have a .beta.-amino alcohol structure.
In the present scientific climate, all stereoisomers of a potential pharmaceutical must be evaluated individually. Consequently, methods of preparatively separating .beta.-blocker stereoisomers and for ascertaining their stereochemical purity are of considerable current interest. Moreover, much effort continues to be expended by pharmacologists in the study of how .beta.-blocker stereochemistry influences the extent and mode of their action. There are now a variety of liquid chromatographic methods which facilitate determinations of stereochemical purity of .beta.-blockers, studies of differences in the rate of metabolism of their enantiomers and studies of the stereochemical pathways of metabolism. While it is possible and often practical to derivatize enantiomers with a chiral reagent so as to obtain diastereomers which are separable on an achiral column, there are potential disadvantages to this approach. In some instances, the enantiomers of .beta.-blockers have been separated on achiral columns through the use of chiral mobile phase additives as reported by C. Petterson, et al. in J. of Chromatogr., 204, 179-384 (1981) and 407, 217-229 (1987). However, the scope of this method remains undetermined, and it too is disadvantageous in some applications. Instances of derivatization with an achiral reagent prior to enantiomer separation on a column containing chiral stationary phases, CSPs, have been reported. However, the need for derivatization, and in the case of preparative separations--dederivatization, is an obstacle to be avoided if possible. The direct separation of underivatized enantiomers on a CSP is to be preferred but is neither always possible nor feasible.
The object of this invention is to provide a process for the separation of underivatized enantiomers of .beta.-amino alcohol compounds, particularly .beta.-blocker drugs.
Another object of this invention is to provide a process for the direct separation of underivatized enantiomers of .beta.-amino alcohol compounds, particularly .beta.-blocker drugs, by means of a chiral selector.
A further object of this invention is to provide a process for the direct separation of underivatized enantiomers of .beta.-amino alcohol compounds, particularly .beta.-blocker drugs, by means of liquid chromatography employing a chiral selector as a chiral stationary phase (CSP).
Yet another object of this invention is to provide a process for the direct separation of underivatized enantiomers of .beta.-amino alcohol compounds, particularly .beta.-blocker drugs, by means of a liquid membrane containing a chiral selector passing in contact with one side of a semi-permeable membrane while a mixture of the enantiomers are in contact with the other side of the semi-permeable membrane.